PTCH1 is the receptor protein of Hedgehog signaling pathway, and Hedgehog pathway plays a vital role in mammalian embryonic development. However, the specific biological role of PTCH1 is incompletely understood for embryonic development. Here, we used a CRISPR/Cas9 genome editing approach to generate a homozygous PTCH1 knock-out iPSC line (SCMCi001-A-1) from a healthy donor, which will be a valuable in vitro model to study the pathogenic mechanism of PTCH1 dysfunction in congenital disease.